The study aims to use a combination of in vitro and in vivo approaches to study the genetic regulation of cell fate choices in the vertebrate spinal chord. The focus of this study is on a diverse population of cells between the motor neurons ventrally and the secondary sensory neurons dorsally that together compromise the interneurons of the spinal chord. The applicant has already defined distinct subpopulations of these post- mitotic cells based on the combinatorial expression of three putative transcription factors, Pax2, Evx1 and En1. Medial to these neurons, in the proliferating ventricular zone, other subpopulations can be defined using different transcription factors, such as Pax3, Pax6, Pax7, ChoxE. This proposal is testing the hypothesis that these medial cell groups are the direct precursors to the lateral neurons, and that the transcription factors expressed by the medial group will control (either directly or indirectly) the unique molecular phenotypes seen in post-mitotic subpopulations. The applicant proposes to try to manipulate the molecular phenotypes of the neuronal pool by treating cultured explants with factors (such as Sonic hedgehog and BMPs) that are known to influence cell fate choices in the presumed precursor pool. Another experimental approach will to overexpress one transcription factor, Pax6, IN THE Precursor pool and see whether this can bias neuronal cell fate choice.